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Journal of Genetic Engineering and Biotechnology, volume 19, issue 1, publication number 155

Expression of acid cleavable Asp-Pro linked multimeric AFP peptide in E. coli

Mollaev Murad 1, 2, 3
Zabolotskii Artur 2, 4
Gorokhovets Neonila 5
Nikolskaya Elena 2, 6
Sokol Maria 2, 6
Tsedilin Andrey 7
Mollaeva Mariia 2, 6
Kuvaev Timofey 8
Pshenichnikova Anna 3
Publication typeJournal Article
Publication date2021-10-14
Quartile SCImago
Q2
Quartile WOS
Impact factor3.3
ISSN1687157X, 20905920
Genetics
Biotechnology
Abstract
Background

Difficult to express peptides are usually produced by co-expression with fusion partners. In this case, a significant mass part of the recombinant product falls on the subsequently removed fusion partner. On the other hand, multimerization of peptides is known to improve its proteolytic stability in E. coli due to the inclusion of body formation, which is sequence specific. Thereby, the peptide itself may serve as a fusion partner and one may produce more than one mole of the desired product per mole of fusion protein. This paper proposes a method for multimeric production of a human alpha-fetoprotein fragment with optimized multimer design and processing. This fragment may further find its application in the cytotoxic drug delivery field or as an inhibitor of endogenous alpha-fetoprotein.

Results

Multimerization of the extended alpha-fetoprotein receptor-binding peptide improved its stability in E. coli, and pentamer was found to be the largest stable with the highest expression level. As high as 10 aspartate-proline bonds used to separate peptide repeats were easily hydrolyzed in optimized formic acid-based conditions with 100% multimer conversion. The major product was represented by unaltered functional alpha-fetoprotein fragment while most side-products were its formyl-Pro, formyl-Tyr, and formyl-Lys derivatives. Single-step semi-preparative RP-HPLC was enough to separate unaltered peptide from the hydrolysis mixture.

Conclusions

A recombinant peptide derived from human alpha-fetoprotein can be produced via multimerization with subsequent formic acid hydrolysis and RP-HPLC purification. The reported procedure is characterized by the lower reagent cost in comparison with enzymatic hydrolysis of peptide fusions and solid-phase synthesis. This method may be adopted for different peptide expression, especially with low amino and hydroxy side chain content.

Citations by journals

1
International Journal of Molecular Sciences
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Citations by publishers

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Multidisciplinary Digital Publishing Institute (MDPI)
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Mollaev M. et al. Expression of acid cleavable Asp-Pro linked multimeric AFP peptide in E. coli // Journal of Genetic Engineering and Biotechnology. 2021. Vol. 19. No. 1. 155
GOST all authors (up to 50) Copy
Mollaev M., Zabolotskii A., Gorokhovets N., Nikolskaya E., Sokol M., Tsedilin A., Mollaeva M., Chirkina M., Kuvaev T., Pshenichnikova A., Yabbarov N. G. Expression of acid cleavable Asp-Pro linked multimeric AFP peptide in E. coli // Journal of Genetic Engineering and Biotechnology. 2021. Vol. 19. No. 1. 155
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TY - JOUR
DO - 10.1186/s43141-021-00265-5
UR - https://doi.org/10.1186%2Fs43141-021-00265-5
TI - Expression of acid cleavable Asp-Pro linked multimeric AFP peptide in E. coli
T2 - Journal of Genetic Engineering and Biotechnology
AU - Mollaev, Murad
AU - Zabolotskii, Artur
AU - Gorokhovets, Neonila
AU - Nikolskaya, Elena
AU - Sokol, Maria
AU - Tsedilin, Andrey
AU - Chirkina, Margarita
AU - Kuvaev, Timofey
AU - Pshenichnikova, Anna
AU - Yabbarov, Nikita G
AU - Mollaeva, Mariia
PY - 2021
DA - 2021/10/14 00:00:00
PB - Springer Nature
IS - 1
VL - 19
PMID - 34648110
SN - 1687-157X
SN - 2090-5920
ER -
BibTex
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BibTex Copy
@article{2021_Mollaev
author = {Murad Mollaev and Artur Zabolotskii and Neonila Gorokhovets and Elena Nikolskaya and Maria Sokol and Andrey Tsedilin and Margarita Chirkina and Timofey Kuvaev and Anna Pshenichnikova and Nikita G Yabbarov and Mariia Mollaeva},
title = {Expression of acid cleavable Asp-Pro linked multimeric AFP peptide in E. coli},
journal = {Journal of Genetic Engineering and Biotechnology},
year = {2021},
volume = {19},
publisher = {Springer Nature},
month = {oct},
url = {https://doi.org/10.1186%2Fs43141-021-00265-5},
number = {1},
doi = {10.1186/s43141-021-00265-5}
}
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